Adaptation to ER stress is mediated by differential stabilities of pro-survival and pro-apoptotic mRNAs and proteins.

The accumulation of unfolded proteins in the endoplasmic reticulum (ER) activates a signaling cascade known as the unfolded protein response (UPR). Although activation of the UPR is well described, there is little sense of how the response, which initiates both apoptotic and adaptive pathways, can selectively allow for adaptation. Here we describe the reconstitution of an adaptive ER stress response in a cell culture system. Monitoring the activation and maintenance of representative UPR gene expression pathways that facilitate either adaptation or apoptosis, we demonstrate that mild ER stress activates all UPR sensors. However, survival is favored during mild stress as a consequence of the intrinsic instabilities of mRNAs and proteins that promote apoptosis compared to those that facilitate protein folding and adaptation. As a consequence, the expression of apoptotic proteins is short-lived as cells adapt to stress. We provide evidence that the selective persistence of ER chaperone expression is also applicable to at least one instance of genetic ER stress. This work provides new insight into how a stress response pathway can be structured to allow cells to avert death as they adapt. It underscores the contribution of posttranscriptional and posttranslational mechanisms in influencing this outcome.

Persistent glycoprotein misfolding activates the glucosidase II/UGT1-driven calnexin cycle to delay aggregation and loss of folding competence.

The UDP-glucose:glycoprotein glucosyltransferase (UGT) is a central player of glycoprotein quality control in the endoplasmic reticulum (ER). UGT reglucosylation of nonnative glycopolypeptides prevents their release from the calnexin cycle and secretion. Here, we compared the fate of a glycoprotein with a reversible, temperature-dependent folding defect in cells with and without UGT1. Upon persistent misfolding, tsO45 G was slowly released from calnexin and entered a second level of retention-based ER quality control by forming BiP/GRP78-associated disulfide-bonded aggregates. This correlated with loss in the ability to correct misfolding. Deletion of UGT1 did not affect the stringency of ER quality control. Rather, it accelerated release from calnexin and transfer to the second ER quality control level, but it did so after an unexpectedly long lag, showing that cycling in the calnexin chaperone system is not frenetic, as claimed by existing models, and is fully activated only upon persistent glycoprotein misfolding.

The noncatalytic portion of human UDP-glucose: glycoprotein glucosyltransferase I confers UDP-glucose binding and transferase function to the catalytic domain.

The eukaryotic cell monitors the fidelity of protein folding in the endoplasmic reticulum and only permits properly folded and/or assembled proteins to transit to the Golgi compartment in a process termed "quality control." An endoplasmic reticulum (ER) lumenal sensor for quality control is the UDP-glucose:glycoprotein glucosyltransferase that targets unfolded glycoproteins for transient, calcium-dependent glucosylation. This modification mediates glycoprotein interaction with the folding machinery comprised of calnexin or calreticulin in conjunction with ERp57. Two human UGT homologues, HUGT1 and HUGT2, exist that share 55% identity. The highest degree of identity resides in the COOH-terminal 20% of these proteins, the putative catalytic domain of HUGT1. However, only HUGT1 displays the expected functional activity. The contribution of the NH2-terminal remainder of HUGT1 to glucosyltransferase function is presently unknown. In this report we demonstrate that HUGT2 is localized to the ER in a manner that overlaps the distribution of HUGT1. Analysis of a series of HUGT1 and HUGT2 chimeric proteins demonstrated that the carboxyl-terminal region of HUGT2 contains a catalytic domain that is functional in place of the analogous portion of HUGT1. Whereas neither catalytic domain displayed detectable activity when expressed alone, co-expression of either catalytic domain with the noncatalytic amino-terminal portion of HUGT1 conferred UDP-Glc binding and transfer of glucose that was specific for unfolded glycoprotein substrates. The results indicate that the amino-terminal 80% of HUGT1 is required for activation of the catalytic domain, whereas the homologous portion of HUGT2 cannot provide this function.

The unfolded protein response in nutrient sensing and differentiation.

Eukaryotic cells coordinate protein-folding reactions in the endoplasmic reticulum with gene expression in the nucleus and messenger RNA translation in the cytoplasm. As the rate of protein synthesis increases, protein folding can be compromised, so cells have evolved signal-transduction pathways that control transcription and translation -- the 'unfolded protein response'. Recent studies indicate that these pathways also coordinate rates of protein synthesis with nutrient and energy stores, and regulate cell differentiation to survive nutrient-limiting conditions or to produce large amounts of secreted products such as hormones, antibodies or growth factors.